We propose to continuously improve the Yale/NIDA Neuroproteomics Center that brings exceptionally strong Yale programs in proteomics and signal transduction in the brain together with neuroscientists from 8 other institutions across the U.S. to create a national resource that will collaborate to identify adaptive changes in protein signaling that occur in response to substances of abuse. Twenty-three faculty with established records of highly innovative research into the molecular actions of psychoactive addictive drugs, as well as of other basic aspects of neurobiology, will work together in a unique synergy with the Keck Foundation Biotechnology Laboratory to create the Center, whose theme is ?Proteomics of Altered Signaling in Addiction?. The Center will use cutting edge proteomic technologies to analyze neuronal signal transduction mechanisms and the adaptive changes in these processes that occur in response to drugs of abuse. With Co-Directors Angus Nairn (Psychiatry) and Kenneth Williams (Mol. Biophys. & Biochem.) in the Administrative Core, the Center includes Discovery Proteomics (DPC) and Targeted Proteomics (TPC) Cores. Biophysical technologies from the DPC will extend protein profiling analyses into the functional domain while lipid analyses from the DPC will leverage proteome level analyses to provide an increasingly biological systems level approach. A Biostatistics and Bioinformatics Core (BBC) that includes high performance computing and the Bioinformatics Support Program in the Yale Medical Library will provide essential support that will leverage the value of each of the proteomic technology cores. A Pilot Research Project Core is a cornerstone in our efforts to encourage strong mentoring relationships that will help attract and train future outstanding scientists. Behavioral adaptations that accompany drug addiction are believed to result from both short and long-term adaptive changes in brain reward centers. Thus, exposure to drugs of abuse regulates intracellular signaling processes that alter gene expression, protein translation, and protein post-translational modifications. Repeated exposure to drugs of abuse leads to stable alterations in these signaling systems that are critical for the changes in brain chemistry and structure of the addicted brain. The Center's research goals include analysis of the actions of cannabis, cocaine, nicotine, and opioids on these intracellular signaling pathways in brain reward areas and development of methods that enable proteomic analysis of the single types of neurons that define the circuits that underlie the actions and addictive properties of drugs of abuse. Targeted and data- independent MS analyses of signaling proteins implicated in the actions of drugs of abuse will be used to analyze the impact of substance abuse on the neuroproteome with motif-based, ?Middle-down? MS/MS, and other novel approaches being used to study protein post-translational modifications and to uncover the interactomes of key proteins. A major initiative of the BBC will be to develop novel methods for deep integration of genomic, transcriptomic, and proteomic data with brain region, cell type and allele-specificity.